The indefinite persistence of virus reservoirs, with the capacity to fuel virus rebound, is the major obstacle to eliminating HIV infection. Identifying reservoirs of HIV-1 and sanctuaries where HIV-1 replication is not completely suppressed by antiretroviral (ARV) therapy (ART) will facilitate the exploration of new approaches to improve treatment outcomes, and will be critical to future efforts to eliminate HIV-1 from the body. Despite extensive study, the location and mechanisms for retention of HIV reservoirs and sanctuaries in the ARV treated host remain poorly understood. Historically, efforts to understand virus reservoirs have centered on easily accessible tissue sites such as the systemic circulation, superficial lymph nodes, and genital fluids. More recently, a few small studies have attempted to examine other sites such as Gut-associated lymphoid tissues (GALT) in the large and small bowel, bronchioalveolar lavage, CSF and kidney. However, we and others have demonstrated that latently infected cells within the peripheral blood and lymph nodes are not significant sources of rebounding virus once therapy is withdrawn or fails, and the origins of these rebounding virus populations remain unclear. A comprehensive investigation of virus reservoirs within the infected host is therefore urgently needed. To this end, we have developed innovative computational tools for the identification of virus reservoirs and compartments from viral gene sequence data. The ideal source of materials for identifying reservoirs, and for assessing mechanisms of retention of reservoirs, would be an invasive examination of both GALT (where >65% of all T-lymphocytes are localized, and the major site of virus replication early in HIV-1 infection) and autopsy-derived tissues from individuals who died while on suppressive ART. Presently, these research areas remain virtually unexplored due to the difficulty in finding subjects that fit these criteria. However, we have identified and obtained tissues from several individuals meeting our criteria and have access to many more autopsies of individuals forming the control group that have died of AIDS while not on ART. Here we propose to systematically explore the entire body for latent viral reservoirs and sanctuaries of active viral replication during ART. We propose to derive HIV-1 RNA and DNA sequences from GALT during suppressive ART, and autopsy tissues from individuals who died while on or not on suppressive ART, appraise the sequences for features of viral reservoirs, and identify tissues with features of sanctuaries of ongoing viral replication. PUBLIC HEALTH RELEVANCE: The indefinite persistence of virus reservoirs, with the capacity to fuel virus rebound, is the major obstacle to eliminating HIV infection. Identifying reservoirs of HIV-1 and sanctuaries where HIV-1 replication is not completely suppressed by antiretroviral (ARV) therapy (ART) will facilitate the exploration of new approaches to improve treatment outcomes, and will be critical to future efforts to eliminate HIV-1 from the body. Here we propose to systematically explore the entire body for latent viral reservoirs and sanctuaries of active viral replication during ART.